Anionic trypsin-like enzymes from the crab Eriocheir japonicus De Haan active in more acidic media

Abstract

Abstract 1. 1. Proteolytic enzymes from digestive fluid of the catadromous crab Eriocheir japonicus De Haan have been investigated. 2. 2. Four types of the enzyme referred to as A 1 , A 2 , A 3 and A 4 are purified by gel filtration and DEAE-Sephadex column chromatography in a homogeneous state (only A 2 contaminated with minor component). 3. 3. Molecular weight of A 2 , A 3 and A 4 is approx. 29,000 on 0.1% SDS-polyacrylamide gel electrophoresis. 4. 4. These enzymes show optimal pH in acidic and neutral media (A 1 , at pH 5.8; A 2 , at pH 6.3; A 3 , at pH 7.0; A 4 , at pH 7.5) and high stability in more alkaline media at pH 5 or above. 5. 5. A 1 is inhibited with phenylmethylsulfonyl fluoride, diisopropylphosphoryl fluoride, and soy bean trypsin inhibitor. 6. 6. A 2 , A 3 and A 4 are inhibited with tosyl- l -lysine chloromethyl ketone, diisopropylphosphoryl fluoride, leupeptin, soy bean trypsin inhibitor and potato protease inhibitor II-a and II-b. 7. 7. A 2 , A 3 and A 4 act on synthetic lysyl and arginyl derivatives, but not on aromatic amino acid derivatives. 8. 8. From the findings including electrophoretic behavior, A 2 , A 3 and A 4 are anionic trypsin-like enzyme and different from A 1 .